Conserved nuclear receptor antagonism balances Hippo and Notch to control tissue growth and tumor suppression

Nuclear receptors orchestrate diverse biological processes through ligand-dependent transcriptional control, yet how competing family members balance oncogenic signaling to maintain tissue homeostasis remains poorly understood. Here, we uncover an evolutionarily conserved antagonism between nuclear receptors that directly controls tumor suppression through coordinated repression of the Hippo and Notch pathways. We identify Hr3 in Drosophila and its mammalian ortholog RORA as potent tumor suppressors that functionally oppose the oncogenic nuclear receptor E75/NR1D2. Using Drosophila genetic models and MARCM clonal analysis, we demonstrate that Hr3 suppresses tissue overgrowth, blocks invasion, and promotes apoptosis across multiple neoplastic contexts. Genome-wide CUT&Tag profiling reveals that Hr3 directly occupies regulatory elements of key Hippo and Notch target genes, with a remarkable 50% overlap with E75 binding sites, suggesting competitive antagonism. Mechanistically, Hr3 physically associates with core transcriptional machinery of both pathways, including Yorkie, Scalloped, Suppressor of Hairless, and Notch intracellular domain (NICD), to repress oncogenic gene expression programs and counteract E75-driven activation. Moreover, this regulatory axis is conserved in human cancer, where RORA overexpression suppresses glioblastoma cell proliferation, migration, and colony formation in a similar mechanism. Our findings establish nuclear receptor competition as a fundamental mechanism governing growth control and reveal therapeutic opportunities for targeting oncogenic transcriptional networks through endogenous regulatory circuits.